Evaluation of a Commercial PCR Kit for Diagnosis of Cytomegalovirus Infection of the Central Nervous System

We evaluated the AMPLICOR cytomegalovirus (CMV) PCR kit for the diagnosis of neurologic CMV infections on 43 positive and 112 negative archived cerebrospinal fluid specimens originally tested by an in-house PCR method. The AMPLICOR kit showed sensitivity and specificity of 95 and 100%, respectively, versus the home-grown assay, indicating its utility in this clinical setting.     

Anticipatory control of head posture.

OBJECTIVE: We tested a hypothesis on two patterns of anticipatory postural adjustments (APAs) in neck muscles, reciprocal and co-activation, that may be used in a task-specific way. We also explored possible relation of APAs in leg and trunk muscles to head stabilization. METHODS: Load perturbations (loading and unloading) were applied to the head, trunk, and head and trunk simultaneously using similar hand actions by standing persons. Electromyographic signals (EMGs) from 10 muscles were recorded. Shifts of the center of pressure and EMG indices were computed over typical time intervals for APA. RESULTS: Time-shifted (reciprocal) activation of neck flexor and extensor muscles during APAs was seen when perturbations were applied directly to the head. Simultaneous activation dominated when the perturbations were applied to the trunk. Minimal APAs were seen in the leg/trunk muscles during head perturbation tests. APAs during trunk perturbation were not different from those during trunk and head perturbation. CONCLUSIONS: The results confirm the existence of two different patterns of APAs in neck muscles. A time-shifted (reciprocal) pattern is more likely to be used in anticipation of a perturbation acting directly on the head. A simultaneous activation (co-activation) pattern is used when direction of head perturbation cannot be predicted with certainty. Leg/trunk APAs are unlikely to help stabilize head posture. SIGNIFICANCE: These results are important for better understanding of feed-forward mechanisms of the control of head posture with possible implications for neurological patients who suffer from impaired feed-forward postural control.     

Freeze-drying polymeric colloidal suspensions: nanocapsules, nanospheres and nanodispersion. A comparative study.

Different polymeric nanoparticles were freeze-dried and the powders compared to determine the influence of the lipophilic core (Miglyol 810) or benzyl benzoate) and polymeric material (poly(epsilon-caprolactone) or Eudragit S90) on their drug content and morphology. Diclofenac, a non-steroidal anti-inflammatory drug, was used as a model. To characterize the products, a biological experiment based on the evaluation of the mucosal toxicity of diclofenac was conducted. Nanocapsule and nanosphere suspensions were prepared by nanoprecipitation and freeze-dried after the addition of colloidal silicon dioxide. The powders were examined under scanning electron microscopy (SEM) and gastrointestinal tolerance of products was evaluated in rats. Powders presented drug contents between 90.2+/-5.5 and 101.1+/-1.9% (HPLC). SEM analyzes showed non-spherical microparticles and, at higher magnifications, the micro-powder surface presented a homogeneous nanocovering. Regarding the gastrointestinal tolerance, with the exception of benzyl benzoate-loaded formulations, powders presented lesional indexes lower than the diclofenac salt solution. In contrast to the literature, nanocapsules can be dried by freeze-drying without leakage of drug or breaking the capsule wall.     

Up-Regulation of TH2 feto-placental levels could be involved in the protective effect of low-molecular-weight heparin treatment on spontaneous abortion in mice

The cytokines that predominate at the healthy maternal fetal interface are compatible with those produced by Th2/3 cells. Th1 and Th2 type immunity are mutually inhibitory and cytokine profiles are regulated in part by maternal sex steroids. The immune and endocrine equilibrium required for pregnancy success may be modified by external factors including stress, infection and altered maternal nutrition. The latter has received surprisingly little attention particularly as the effects of nutrition on immunity per se are widely documented. We have used animal models to investigate the effects of altered maternal diet on both immune and endocrine mechanisms important for pregnancy success. In rodents, maternal deficiencies in iron and vitamin A have been shown to negatively alter the expression of placental cytokines and in the case of vitamin A, increase placental apoptosis. In a highly controlled sheep model, overfeeding young growing females carrying singleton pregnancies restricts placental growth resulting in the premature delivery of low birth weight lambs. This is associated with reduced maternal concentrations of progesterone, placental lactogen, PSPB, GH and increased insulin, IGF-1, leptin and thyroid hormones (Wallace et al. Reprod 2001; 122:347–357). At day 80 of gestation (term=145) the placentae of overfed dams exhibit reduced expression of proliferation markers, predominantly in the fetal trophectoderm, and increased expression of the pro-apoptotic protein bax. These data indicate an altered balance between placental proliferation and apoptosis, possibly linked to maternal endocrine status. We conclude that maternal diet has considerable impact on immuno-endocrine mechanisms critical for pregnancy success.     

A Decline in C6 Antibody Titer Occurs in Successfully Treated Patients with Culture-Confirmed Early Localized or Early Disseminated Lyme Borreliosis

C₆, a Borrelia burgdorferi-derived peptide, is used as the antigen in the C₆-Lyme disease diagnostic test. We assessed retrospectively whether a fourfold decrease or a decrease to a negative value in anti-C₆ antibody titer is positively correlated with a positive response to treatment in a sample of culture-confirmed patients with either early localized (single erythema migrans [EM]; n = 93) or early disseminated (multiple EM; n = 27) disease. All of these patients had been treated with antibiotics and were free of disease within 6 to 12 months of follow-up. Results show that a serum specimen taken at this time was either C₆ negative or had a ≥4-fold decrease in C₆ antibody titer with respect to a specimen taken at baseline (or during the early convalescent period if the baseline specimen was C₆ negative) for all of the multiple-EM patients (P < 0.0001) and in 89% of the single-EM patients (P < 0.0001). These results indicate that a decline in anti-C₆ antibody titer coincides with effective antimicrobial therapy in patients with early localized or early disseminated Lyme borreliosis.     

Radiation-induced lung injury in vivo: Expression of transforming growth factor—Beta precedes fibrosis

Cytokine release from irradiated cells has been postulated to start soon after irradiation preceding detectable clinical and pathological manifestation of lung injury. The expression of transforming growth factor beta (TGF), a fibrogenic and radiation-inducible cytokine, was studied from 1–16 weeks after the 15 and 30 Gray (Gy) of thoracic irradiation to rats. Thoracic irradiation caused an increase in TGF protein in bronchoalveolar lavage (BAL) fluid peaking at 3–6 weeks as compared to sham-irradiated control rats. Steady state TGF mRNA expression as shown by whole lung northern blot assay paralleled the TGF protein expression in BAL fluid. The peak of TGF protein increase in BAL fluid between 3 and 6 weeks coincided with the initial influx of inflammatory cells in BAL fluid, but preceded histologically discernable pulmonary fibrosis that was not apparent until 8–10 weeks after irradiation. In conclusion, TGF and mRNA and protein upregulation preceded the radiation-induced pulmonary fibrosis, suggesting a pathogenetic role in the development of radiation fibrosis.     

PCR analysis of egyptian respiratory adenovirus isolates, including identification of species, serotypes, and coinfections

Eighty-eight adenovirus (Ad) isolates and associated clinical data were collected from walk-in patients with influenza-like illness in Egypt during routine influenza surveillance from 1999 through 2002. Respiratory Ad distributions are geographically variable, and serotype prevalence has not been previously characterized in this region. Serotype identity is clinically relevant because it predicts vaccine efficacy and correlates strongly with both clinical presentation and epidemiological pattern. Species and serotype identities were determined using several well-validated multiplex PCR protocols culled from the literature and supplemented with a few novel primer sets designed to identify rare types. The isolates included common species B1 serotypes (Ad3 and Ad7), common species C serotypes (Ad1, Ad2, and Ad5), the less common species B2 serotype Ad11, and three isolates of the rare species B1 serotype Ad16. Two isolates that appear to be variant Ad16 were also identified. Fifteen coinfections of multiple adenoviral types, primarily AdB/AdC and Ad3/Ad7 dual infections, were detected. The majority of these were verified using redundant PCR tests targeted at multiple genes. PCR is able to resolve coinfections, in contrast to traditional serum neutralization tests. PCR is also comparatively rapid and requires very little equipment. Application of the method allowed an inclusive determination of the serotypes found in the Egyptian respiratory sample set and demonstrated that coinfections are common and may play a previously unrecognized role in adenovirus pathogenesis, evolution, and epidemiology. In particular, coinfections may influence adenoviral evolution, as interserotypic recombination has been identified as a source of emerging strains.     

Epidemiology of Cyclospora cayetanensis and other intestinal parasites in a community in Haiti

We conducted an exploratory investigation in a community in Haiti to determine the prevalence of Cyclospora cayetanensis infection and to identify potential risk factors for C. cayetanensis infection. In 2001, two cross-sectional stool surveys and a nested case-control study were conducted. In 2002, a follow-up cross-sectional stool survey was conducted among children < or =10 years of age. Stool specimens from study participants and water samples from their wells were examined for Cyclospora and other intestinal parasites. In stools, the prevalence of infection with Cyclospora in persons of all ages decreased from 12% (20 of 167 persons) in February 2001 to 1.1% (4 of 352 persons) in April 2001, a 90.8% decrease. For children < or =10 years of age, the prevalence rates were 22.5% (16 of 71 children) in February 2001, 3.0% (4 of 135 children) in April 2001, and 2.5% (2 of 81 children) in January 2002. Use of the water from the artesian well in the northern region of the community versus the one in the south was the only risk factor associated with Cyclospora infection in multivariate analyses (odds ratio, 18.5; 95% confidence interval, 2.4 to 143.1). The water sample from one of the nine wells or water sources tested (one sample per source) in January 2001, shortly before the investigation began, was positive for Cyclospora by UV fluorescence microscopy and PCR. None of the water samples from the 46 wells or water sources tested during the investigation (one sample per source per testing period, including the artesian wells) were positive for Cyclospora. Further studies are needed to assess the role of water as a possible risk factor for Cyclospora infection in Haiti and other developing countries.     

Pemphigus Vulgaris Autoantibody Response is Linked to HLA-DQB10503 in Pakistani Patients

ABSTRACT: Pemphigus vulgaris (PV) is an autoimmune disease of the skin and mucous membranes characterized by an autoantibody response against an epidermal cadherin. We performed high resolution HLA class II typing in 19 patients with PV from Rawalpindi, Pakistan and 19 non-Jewish European PV patients from Boston by sequence-specific oligonucleotide probe hybridization. The results were compared with two separate ethnically matched control populations. We found that PV patients from Pakistan had significantly increased frequencies of DRB1*1404 ( p = 0.01), DQA1*0101 ( p = 0.02), and DQB1*0503 ( p = 0.01). Among the patients of non-Jewish European ancestry, DRB1*1401 ( p < 10⁻⁶), DQA1*0101 ( p < 10⁻⁵) and DQB1*0503 ( p < 10⁻⁶), were increased in PV patients. Formal linkage analysis between the major histocompatibility complex and the PV antibody was performed in 67 relatives of the 19 Pakistani patients. The results showed strong evidence for linkage of HLA-DRB1*1404, DQA1*0101, DQB1*0503, with the presence of PV antibody in relatives’ families with a significant logarithm of the odds score of 6.06. Based on the three dimensional structure of class II molecules, we propose that HLA-DQA1*0101 and DQB1*0503, encode a negatively charged P9 peptide binding pocket of the DQ molecule and are significantly associated with susceptibility to PV in non-Jewish populations.     

Five-test simple scheme for species-level identification of clinically significant coagulase-negative staphylococci

A working scheme developed in our laboratory for identification (by species group and species) of coagulase-negative staphylococci (CNS) was evaluated with 201 consecutive isolates and then validated by using the reference method of Kloos and Schleifer (W. E. Kloos and K. H. Schleifer, J. Clin. Microbiol. 1:82-88, 1975). This five-test simple scheme (referred to here as the simple scheme) combines the novobiocin susceptibility test with tests for urease, pyrrolidonyl arylamidase, ornithine decarboxylase, and aerobic acid from mannose. The addition of one or two tests within a particular species group could then positively identify the isolate. Two commercial systems, Staph-Zym (Rosco) and API-Staph (bioMérieux), along with results obtained by using Rosco diagnostic tablets (nongrowth tests), were also compared with the reference method. One isolate could not be identified even by the reference method. Of the remaining 200 strains, 191 (95.5%) strains were correctly identified with Staph-Zym and 171 strains (85.5%) were correctly identified with API-Staph. The most frequent clinical CNS species isolated were Staphylococcus epidermidis (50.5%), S. haemolyticus (18.5%), S. saprophyticus subsp. saprophyticus (16.0%), S. lugdunensis (6.0%), and S. warneri (2.5%). The simple scheme validated with the reference method has demonstrated an excellent correlation in the identification of the three most frequent species isolated: S. epidermidis, S. haemolyticus, and S. saprophyticus subsp. saprophyticus. With the simple scheme, identification of CNS was possible within 24 h after the enzymatic tests were used, whereas up to 72 h is necessary for the growth tests. This methodology would be very useful in any clinical microbiology laboratory for the presumptive identification of CNS species groups and species.